Fig. 3: Induced mitochondrial fission by localized recruitment of FKBP-BcPI-PLC3A is associated with the rapid disassembly of OMM-associated Drp1 puncta.
a Representative images of HEK293A cells (5 μm scale bar) showing the localization of the OMM-targeted FRB recruiter (OMM-FRB-mRFP, magenta) and Drp1 (mNGHO-Drp1, green) before (top row panels) or 10 min after (bottom row panels) rapamycin-induced (100 nM) recruitment of the catalytically active BcPI-PLC3A (emiRFP670-FKBP-BcPI-PLC3A, gray) to the cytosolic membrane leaflet of the mitochondria. The early phase of the bulk fragmentation response of the mitochondrial network is shown over the initial 300 sec after rapamycin treatment in the inset panels (2.5 μm scale bar). b Representative composite images of COS-7 cells (5 μm scale bar) showing localization of the OMM-targeted FRB recruiter (OMM-FRB-ECFP, green) and Drp1 (mCherry-Drp1, magenta) in response to rapamycin-induced (100 nM) recruitment of the catalytically active FKBP-BcPI-PLC3A to the cytosolic membrane leaflet of the mitochondria. The bulk fragmentation response of the mitochondrial network is presented over the 150 sec period immediately after rapamycin treatment, while a detailed time series of the initial 45 sec post-rapamycin treatment (lower inset panels; 5 μm width); which is the time frame consistently associated with the majority of the OMM fission events, are presented separately as inset image panels. Image series presenting the individual image channels and overlays of the OMM-FRB together with either the NES-MmPKDC1a,b probe, or Drp1 are provided in Supplementary Movie 2.
